Swarming is a specialized form of bacterial motility that develops when cells that can swim are grown in a rich medium on a moist surface, e.g., agar. The cells become multinucleate, elongate, synthesize large numbers of flagella, and advance across the surface in a thin layer of fluid. The fluid plays a significant role in the physiology of swarming by providing the microenvironment in which cells can swim and interact. Swarming promotes invasiveness of infectious pathogens and thus is medically relevant. With E. coli, the model organism that we study, the cells actually swim in a thin film of fluid between two fixed surfaces, a surfactant monolayer above (pinned at its edges) and agar below. Ahead of the swarm, fluid streams clockwise (with the swarm viewed from above) driven by the motion of flagella of cells stuck to the agar near the swarm edge. Within the body of the swarm, fluid drifts both inwards and outwards toward a region about 100 m from the swarm edge. This is a region of rapid cell growth and high surface cell density. For cells to grow and the swarm to expand, fluid must be extracted from the underlying agar. A long-standing but untested hypothesis is that the swarm fluid is drawn from the substrate by osmotic flow. We propose to test this hypothesis by measuring the spatial-temporal dynamics of changes in osmolarity in the swarm fluid near the swarm edge. We will measure changes in the size of liposomes filled with self-quenching and non-self- quenching dyes (the latter as a control) in a medium containing colloidal silica, added to prevent liposomes from sinking and being entrapped by the agar. This project is focused on a major unanswered question central to swarm dynamics, but involves methods that can be applied, as well, to real-time measurements of other important physiological parameters involved in swarm-cell differentiation. We also will try to develop buoyant particulate probes for viscosity and pH. PUBLIC HEALTH RELEVANCE: We are trying to understand how flagellated bacteria sense surfaces and establish the microenvironment required for swarming, a process used by pathogenic bacteria to invade and colonize human tissue or medical devices, an important factor in the development of disease. We will develop methods for measurements of key physical parameters within a swarm: osmolarity, viscosity, and pH.